Ischemic preconditioning (PC) is a phenomenon whereby various stimuli, which themselves do not cause neuronal injury, will significantly reduce the vulnerability of the brain to subsequent lethal ischemia. Ischemic tolerance conferred by a PC stimulus appears to require new gene transcription and protein synthesis. Multiple intracellular pathways have been identified that initiate the PC response, but the genes ultimately responsible remain elusive. Microarray analysis permits the rapid identification and semi-quantification of a large number of genes, making it ideal for studying complex mechanisms such as PC. The hypothesis of the proposed work is that microarray analysis will permit the identification of genes upregulated or downregulated by the PC stimuli and ultimately confer ischemic tolerance. Because many genes are modulated following a noxious stimulus, most of which are not involved in PC, the challenge is to separate relevant genes from those that are mere epiphenomena. Pharmacologic blockade of the PC cascade during application of the PC stimulus can help filter relevant from epiphenomena genes. The following are the specific aims of the proposed work: 1) To identify the genes whose transcription is modulated by sublethal ischemia and separate genes relevant to PC from those that are epiphenomena by the use of pharmacologic inhibitors of PC; 2) To identify the genes whose transcription is modulated following a lethal ischemic insult in preconditioned versus non-preconditioned animals; 3) To create a pool of gene candidates with potential involvement in mediating PC; 4) To determine the PC candidate genes and to confirm their modulation by rt-PCR, immunohistochemistry, and Western blotting; 5) To confirm the relevance of the identified genes to PC by pharmacologically modulating the genes in vivo and then to determine the effect of this modulation on neuronal outcome. The ultimate goal of the proposed research is to identify neuroprotective programs endogenous to neurons and to evoke these programs in patients who are at risk for cerebral ischemia. A substantial benefit of the research will be the identification of novel therapeutic targets upon which future drug development can be based.